Liquid chromatography is a method for separating individual compounds in a mixture of compounds. Liquid chromatography is often used for sample analysis or for purification of a particular compound in a sample mixture. Typically, a sample containing a number of compounds is injected into a fluid stream (typically a solvent of constant or varying composition,) and directed through a chromatographic column. The column separates the compounds in the sample mixture in response to their differential retention in the column. Concentration peaks associated with the separated compounds typically emerge in sequence from the column. Sample purification typically entails higher solvent flow rates, and larger sample quantities, than sample analysis.
The chromatographic peaks are often characterized with respect to their retention time, that is, the time in which the center of the band transits the detector relative to the time of injection. In many applications, the retention time of a peak is used to infer the identity of the eluting analyte based upon related analyses incorporating standards or calibrants. The presence of the separated species are often distinguished through use of a refractometer or an absorbtometer utilizing ultraviolet (UV) light.
In addition to separation columns and detectors, liquid-chromatography (LC) instruments typically include solvent reservoirs, pumps, filters, check valves, sample-injection valves, and other fluid-handling and analysis components. Typically, mobile-phase solvents are stored in reservoirs, and delivered via reciprocating-cylinder based pumps. Sample materials are often injected via syringe-type pumps. For example, some LC systems inject a sample by aspirating (pulling) a fluid-based sample into a tube via a needle or capillary and then into a sample loop. The sample is then injected from the sample loop into the mobile-phase stream on its way to a separation column.
In many scientific or industrial applications, compounds are purified for testing, for further analysis, or for volume production. For target purification, it is often desirable to recover the target compound with as high a purity as possible, and to separate the largest possible quantity of a sample with each run to reduce labor, run time, and other costs. Thus, it is often desirable to process samples at high flow rates to obtain purified compounds in larger quantities and/or more quickly. It is also commonly desirable to operate at pressures in the range of 1,000-5,000 pounds per square inch (psi) or higher. Such pressures are encountered in high-performance (also known as high-pressure) liquid chromatography (HPLC).
Processing liquids at high flow rates and high pressures, however, can cause problems. Pump-blended solvents often encounter outgassing, either due to pressure drops experienced in the pump head or solubility reduction upon solvent mixing. Outgassing in the pump head can impair mixing, and outgassed bubbles, in general, can impair LC performance by altering the behavior of various components. For example, bubbles passing through an optical detector can alter the spectral behavior of the detector, e.g., by causing spectral spikes.
As noted, bubble production can occur during solvent mixing due lower gas solubility of some solvent mixtures relative to the unmixed solvents (such as water and methanol.) One solution to this problem, at least for isocratic analysis, is to use premixed solvents.
More general solutions to bubble production include helium sparging and vacuum degassing. Often used to degas pre-mixed solvents, helium sparging is particularly effective where the solvent reservoir can be pressurized. Sparging, however, can be costly and inconvenient. In-line vacuum degassers offer an alternative solution. Such degassers utilize a semipermeable membrane disposed in an evacuation chamber that removes dissolved gases. Degassers, however, also add cost, and can be ineffective for high flow rates as often utilized in preparative- and process-scale liquid chromatography.